Parasitic infection in various stages life of cultured Acipenser persicus.

The present study was conducted to evaluate the status of the parasite fauna in Acipenser persicus at different development stages, in order to find prevention protocols for parasitic diseases in this valuable species. For this purpose, sampling from each sex breeder, 10 egg samples, 5-day-old larvae (n = 20), 20-day-old larvae (n = 80) and fingerling of A. persicus (n = 60) released in earthen ponds were done. After the bioassay and preparing wet mount from the internal and external organs, identification was done according to the keys. According to the results, no fauna parasites were isolated from egg samples and 5-day-old larvae; but Trichodina spp. was isolated from 20-day-old larvae. Also, the same protozoan was isolated from fingerling released in earthen ponds, the mean intensity, prevalence and range of contamination by fingerling were higher with compared to 20-day-old larvae. Trichodina sp. and Diplostomum spathaceum were isolated from skin and eyes of females, respectively. However, Trichodina sp. and Ichthyophthirius multifiliis were isolated from skin of male breeders. In this study, no parasites were isolated from internal organs of larves and fingerling but four intestinal parasites included: Cucullanus sphaerocephlaus, Anisakis sp., Skyrjabinopsilus semiarmatus, and Lepto-rhynchoides plagicephalu were isolated from internal organs of breeder. Based on a wide range of parasitic infection observed in various life stages of A. persicus, it seems necessary to consider hygienic and management measures.


Introduction
Acipenser persicus is one of the most important species in the family of Acipenseridae which its meat and caviar are considered economically. This fish is distributed along by Koura River (Azerbaijan) and Sefidrood River (Iran). 1 During the past two decades, overfishing, loss of habitat for natural reproduction, industrial and domestic pollution and increase of infectious diseases puts the reservoir in alarming rate. 1 Therefore, artificial culture of A. persicus has been increased in aquaculture industry of Iran. In-breeding and rearing sturgeon aquaculture requires comprehensive information about health status and diseases, not only conduce to the promotion of health and quality of sturgeon, but also could have an important role in the production and proliferation of endangered species. 2 Parasitical pathogens could effect on physiological and biological features and their mechanical damages may also predispose fish to the viral, bacterial and fungal disease and cause severe mortality. 3 Therefore, identification of infectious pathogenic parasites could be helpful in proliferation and rearing of different sturgeon species, especially A. persicus.
So far, more than 60 species of parasites in sturgeons of the Black Sea and the Caspian Sea have been recorded. 4  This study was conducted to investigate the parasitic fauna of cultured A. persicus (in propagation and rearing sturgeon centers) in different growth stages of life in northern of Iran.

Materials and Methods
This study was conducted in Shahid Rajaee Propagation and Rearing Center in the north of Iran. The experiments were conducted under identical conditions, following completely randomized design. 8 A total of 5 samples from each sex breeders, 10 egg samples from female breeders, 20 samples from 5-day-old larvae, 80 samples from 20-day-old larvae and 60 samples from fingerling (that released in earthen ponds) were selected randomly. Samples were transferred by plastic bags contained oxygen to central laboratory of the Caspian Sea Institute, Sari, Iran. 8 After the biometric measurements, wet mount from internal and external organs of larvae, skin, fins, gills and eyes of fingerling and breeders were prepared. 10 After the preparation of wet smear, external parasites was identified by Bazari Moghaddam et al. key, using Klein's silver impregnation technique. 9 In order to investigate gastrointestinal parasites, macroscopic examination was done and then the entrails of the fish were removed. Also, approximately 50 to 60 eggs samples were collected from the ovary of female breeders and transferred to the central laboratory of the Caspian Sea Institute.
All collected parasites specimens were removed and stored in 70% alcohol. Then, cestodes, trematodes and acanthocephalans were stained with aqueous acetocarmine and nematodes were cleared in lacto phenol. 11 The specimens were identified by Bauer and Moravec keys. 12,13 Statistical analysis. The data were subjected to statistical analysis using the SPSS (Version 18; SPSS Inc., Chicago, USA). Results of this study were analyzed statistically using One -way analysis of variance (ANOVA) and the significance level was expressed as p < 0.05. Also, Tukey's test applied to compare between different groups. Mean intensity was determined by dividing the total number of recovered parasites to the number of infected fish samples. Prevalence was also calculated by dividing the number of infected fish samples by the total number of examined ones and expressed as a percentage.
In this survey, Anisakis sp. was isolated (Fig. 3) only from intestine of male breeds (10.00%), while others were found in both sex breeders (p < 0.05, r = 0.034). In intestinal infection, Cucullanus sphaerocephlaus had the highest intensity and frequency which, were observed in both examined sex breeders (p > 0.05, r = 0.124).

Discussion
In recent years, due to losses in A. persicus stocks in the Caspian Sea, many researchers have paid attention to revive the sources of this valuable species. 1 By increasing and developments in rearing of sturgeons in ponds of Iran, identification of pathogenic agents in order to apply the best methods of prevention and treatment seemed necessary.
In this study, no parasites were isolated from 5-day-old larvae of A. persicus; but Trichodina sp. was isolated from 20-day-old larvae, these results were similar to Bazari Moghaddam et al. study. This finding may be related to the short larval rearing period in Vniro ponds (varied between 2-3 weeks), absence of intermediate hosts and also the lower water temperature in ponds compared to fingerlings in earthen ponds and lower organic load in Vniro ponds, presence of parasitic agents seemed lower than in rearing in earthen ponds. [9][10][11] 15 Also, Sattari and Khara et al. not only reported these parasites, but also isolated Eustrongylides excicus in sturgeon's species in southwest coast of Caspian Sea. 16,17 Ghoroghi reported seven intestinal parasites in A. persicus and indicated that C. sphaerocephlaus and S. semiarmatus had the highest intensity and contamination rate. 14 Also, Bazari Moghaddam found that C. sphaerocephlaus had the highest intensity and contamination rate between intestenial parasites in A. persicus breeders, 11 these results was similar to our results and confirmed it. The increase in these parasite communities may be related to the occurrence of Nereids (polychaeta), the intermediate host for C. sphaerocephalus. 6 In the present study, Anisakis sp. was only found in male breeders. However, Ali Mohammadi et al. reported this nematoda in both sex of breeders. 15 Sattari and Sattari and Mokhayer isolated Anisakis sp. from other sturgeon species such as: Huso huso and A. persicus. 8,16 By considering the high ability of this nematode to cause disease in humans and creating the allergic reactions, comprehensive studies on Anisakidea and other zoonotic parasites in sturgeons species seems to be essential. [18][19][20] Due to important roles and negative effects of parasites infection in different growth stages of life in sturgeons and based on the results of previous studies, [15][16][17][18] it seems following protocols should be observed; proper pond designing, controlling the water quality and quantity, using of disinfections, controlling of snails and aquatic birds as intermediate hosts, improving nutrition and decreasing stress condition in order to reduce the numbers of parasites infection and improvement on the health conditions of cultured sturgeons. 19 Based on the results, periodic parasitical examination can contribute to the control of fish parasite and a reduction in the economic losses in propagation and rearing center of sturgeons species.